Engineering LmrR protein for L-proline-based asymmetric aldol biocatalysis

Area of Science:

• Biocatalysis
• Protein Engineering
• Organic Chemistry

Background:

• L-proline is a potent organocatalyst for asymmetric synthesis.
• In proteins, proline residues are typically inactive due to peptide bond formation.
• Naturally occurring proline-based enzymes are rare.

Purpose of the Study:

• To engineer a nonenzymatic protein scaffold into a biocatalyst.
• To leverage a native L-proline residue for enantioselective catalysis.
• To develop a new-to-nature L-proline-based enzyme for asymmetric synthesis.

Main Methods:

• Engineering the LmrR protein scaffold.
• Exposing the N-terminal L-proline residue.
• Rational design, protein engineering, and reaction optimization.
• Catalysis of enantioselective aldol reactions.

Main Results:

• Developed an engineered LmrR variant with catalytic activity.
• Achieved up to 99% conversion in aldol reactions.
• Obtained >99% enantiomeric excess with various aldehydes.
• Demonstrated broad substrate scope for aromatic and heteroaromatic aldehydes.

Conclusions:

• A novel strategy to unlock dormant catalytic potential in amino acids and protein scaffolds.
• Created a tailored, L-proline-based enzyme for asymmetric synthesis.
• Provided an applicable approach for designing new biocatalysts.